Eukaryotic DNA contains many origins of replication. At each origin, a multiprotein origin recognition complex binds to initiate the unwinding of the DNA.

Licensing of DNA replication Eukaryotic cells utilize thousands of origins, and so the entire genome can be replicated in a timely manner. The use of multiple origins, however, creates a special problem in the timing of replication: the entire genome must be precisely replicated once and only once in each cell cycle so that no genes are left unrepli-cated and no genes are replicated more than once. How does a cell ensure that replication is initiated at thousands of origins only once per cell cycle?

The precise replication of DNA is accomplished by the separation of the initiation of replication into two distinct steps. In the first step, the origins are licensed, meaning that they are approved for replication. This step is early in the cell cycle when a replication licensing factor attaches to an origin. In the second step, initiator proteins cause the separation of DNA strands and the initiation of replication at each licensed origin. The key is that initiator proteins function only at licensed origins. As the replication forks move away from the origin, the licensing factor is removed, leaving the origin in an unlicensed state, where replication cannot be initiated again until the license is renewed. To ensure that replication takes place only once each cell cycle, the licensing factor is active only after the cell has completed mitosis and before the initiator proteins become active.

Unwinding Several helicases that separate double-stranded DNA have been isolated from eukaryotic cells, as have single-strand-binding proteins and topoisomerases (which have a function equivalent to the DNA gyrase in bacterial cells). These enzymes and proteins are assumed to function in unwinding eukaryotic DNA in much the same way as unwinding in bacterial cells.

Eukaryotic DNA polymerases A significant difference in the processes of bacterial and eukaryotic replication is in the number and functions of DNA polymerases. Eukaryotic cells contain a number of different DNA polymerases that function in replication, recombination, and DNA repair

(Table 12.5). DNA polymerase a, which contains primase activity, initiates nuclear DNA synthesis by synthesizing an RNA primer, followed by a short string of DNA nucleotides. After DNA polymerase a has laid down from 30 to 40 nucleotides, DNA polymerase 8 completes replication on the leading and lagging strands. DNA polymerase ß does not participate in replication but is associated with the repair and recombination of nuclear DNA. DNA polymerase y replicates mitochondrial DNA; a -y-like polymerase also replicates chloroplast DNA. Similar in structure and function to DNA polymerase 8, DNA polymerase e appears to take part in nuclear replication of both the leading and the lagging strands, but its precise role is not yet clear. Other DNA polymerases (£, 0, k, X, allow replication to bypass damaged DNA (called translesion replication) or play a role in DNA repair. Many of the DNA polymerases have multiple roles in replication and DNA repair (see Table 12.5). __

Concepts 9

There are at least thirteeen different DNA polymerases in eukaryotic cells. DNA polymerases a and 8 carry out replication on the leading and lagging strands.


DNA polymerases in eukaryotic cells

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